Implantable materials

ABSTRACT

PCT No. PCT/AU87/00043 Sec. 371 Date Dec. 14, 1987 Sec. 102(e) Date Dec. 14, 1987 PCT Filed Feb. 17, 1987 PCT Pub. No. WO87/05038 PCT Pub. Date Aug. 27, 1987.Polymer hydrogels are adapted for surgical implants by chemical modification of the surface to stimulate the attachment and growth of cells thereto. The modification may be by oxidative acid etching or by copolymerization with methacrylic acid and diethylaminoethyl methacrylate.

TECHNICAL FIELD

This invention relates to implantable materials and to processes fortheir manufacture.

BACKGROUND ART

Blood vessels and naturally occurring internal organs are lined with athin layer of endothelial cells which have a number of bio-chemicalfunctions. In so far as surgical implants are concerned, one importantfunction of endothelial cells is their involvement in the processes ofrendering the surfaces of blood vessels non-thrombogenic.

A key factor in attaining a non-thrombogenic vascular graft is the rapiddevelopment of a lining of the endothelial cells on the implant. Thus,such implants benefit from having surfaces that encourage endothelialcell attachment and spreading.

Similar considerations apply in respect of other implants that areintended for prolonged implantation where blood contact is required suchas permanent indwelling catheters for drug administration, fluiddrainage tubes, vascular shunts, pacemaker leads and implantabletransducers.

Synthetic polymer hydrogels have found a wide range of biomedicalapplications, including controlled drug delivery systems, replacementblood vessels, would dressings, coatings for biosensors, soft tissuesubstitution and contact lenses.

As a family of polymeric materials, synthetic hydrogels are generallywell tolerated when implanted in vivo and can be tailored to suit themany potential functions of prosthetic devices in contact with blood orsoft tissues. The success of hydrogels as biomaterials lies partially intheir superficial resemblance to living tissue, a property attributableto their relatively high water content (say 20%-99%), which immediatelyresults in minimal frictional irritation of surrounding tissues.

In addition, hydrogels can be non-toxic, chemically stable and (due totheir water content) can exhibit a low interfacial tension with aqueousenvironments. This latter property becomes particularly important inconsidering the compatibility of blood-contacting surfaces, whereminimal interfacial tension has been related to thromboresistance.

The hydrogel polyHEMA--poly(2-hydroxyethyl methacrylate)--is known topossess inherent characteristics of good permeability, water uptake andtolerable polymer/tissue interface disruption which make it a desirablebiomaterial (see, for example, Cohn D et al (1984) Radiation--GraftedPolymers for Biomaterial Applications I. 2-Hydroxyethyl Methacrylate:Ethyl Methacrylate Grafting on to Low Density Polyethylene Films J. App.Pol. Sci. 29, 2645-2663).

Despite these advantages, however, unmodified polyHEMA does not have theability to sustain mammalian cell growth and consequently its use as abiomaterial has been restricted to applications where this inability isa positive advantage (see Andrade J. D. (1975) Hydrogels for Medical andRelated Applications ACS SYMPOSIUM SERIES 31, Washington).

Recent investigations into the effects of treating polystyrene withsulphuric acid (see Curtis A.S.G. et al (1983) Adhesion of Cells toPolystyrene Surfaces J. Cell Biol 97, 1500-1506) have shown a markedimprovement in the ability of that polymer to support mammalian cellgrowth after acid etching. Whilst this is not a new concept, modernanalytical methods such as electron spectroscopy for chemical analysishave allowed a more detailed study of surface changes occurring withsuch treatments resulting in some clarification of certain aspects ofcell adhesion.

Another disadvantage of polyHEMA is that its poor mechanical propertiesprevent it from being used as an implant requiring high mechanicalstrength.

By copolymerisation of polyHEMA with other selected synthetic polymers,it has been possible to manipulate surface charges, hydrophilicity andequilibrium water content to achieve varying degrees of attachment andgrowth of fibroblastoid cells. An alternative modification has been toincorporate natural polymers such as collagen, elastin and fibronectinin polyHEMA hydrogels. This has provided a model system to study thecontribution of such extracellular matrix components to cell adhesionand growth. While this approach has allowed the growth of a widervariety of cell types on such hydrogels, it also places otherrestrictions and problems on the polyHEMA system as a biomaterial foruse in prosthetic devices.

It is an object of this invention to provide an implantable materialhaving improved biocompatibility arriving from enhanced endothelial cellattachment properties.

It is a further object of this invention to provide an improvedimplantable material having a mechanically acceptable substrate to whichis attached a polyHEMA layer.

DISCLOSURE OF THE INVENTION

According to the invention there is provided an implantable materialcomprising a hydrogel the surface of which is chemically modified so asto stimulate the attachment and growth of cells thereto.

The chemical modification may consist of hydrolytic etching of thehydrogel or copolymerisation of the hydrogel with methacrylic acid.

The surface of a hydrogel of polyHEMA may also be modified by limitedsurface hydrolysis.

By exposing polyHEMA to a particular acid treatment we have achievedcell attachment and cell growth rates comparable with those of tissueculture polystyrene and better than P.T.F.E (TEFLON), a commonly usedbiomaterial.

The treated hydrogel of the invention has a surface that supportsefficient adhesion of endothelial cells which grow to confluence.Electron spectroscopy chemical analysis of the acid etched polyHEMAindicates an increase in C═O groups relative to C--O groups suggestingthat increased negative charge from carboxyl groups contribute to thechange in cell-substatum interaction. This contention is supported bythe fact that methacrylic acid contributes mainly to increased carboxylgroups in the material.

The cell plating efficiency of an acid etched polyHEMA treated inaccordance with the invention increased from 0% to 95% of that ofglow-discharge treated polystyrene and fibronectin binding capacityincreased from 0 to 2×10⁻¹¹ pico mole per square centimeter.

The invention also provides an implantable material consisting of amechanically acceptable substrate having a hydrogel layer, said hydrogellayer being chemically modified as described above so as to stimulatethe attachment and growth of cells thereto.

The substrate may be any convenient material such as polyurethane,TEFLON, DACRON or other plastic material, platinum, titanium or othermetal as well as carbon and ceramic materials. The polyHEMA may beattached to the substrate by mechanically keying it to a microporousstructure or by grafting in the case of a polymer surface.

According to another aspect of the invention, there is provided a methodof providing an implant comprising the steps of:

(a) forming a substrate having pre-determined mechanical properties,

(b) applying a hydrogel layer to the substrate, and,

(c) chemically modifying the surface of the hydrogel so as to stimulatethe attachment and growth of cells thereto.

BRIEF DESCRIPTION OF THE DRAWINGS

In order that the invention may be more readily understood and put intopractical effect, reference will now be made to the accompanyingdrawings in which:

FIG. 1 is a graph of percentage cell attachment as a function of etchingtime for a polyHEMA substrate etched with sulphuric acid,

FIG. 2 is a graph of B.A.E. cell growth as a function of time forsulphuric acid etched polyHEMA and glow-discharge treated polystyrene,

FIG. 3 is a pictorial representation of the protein bindingcharacteristics of etched and non-etched polyHEMA and rough and smoothP.T.F.E., and,

FIG. 4 is a graph of percentage cell attachment as a function ofpercentage copolymer in polyHEMA after copolymerising charged HEMAmonomer and with methacrylic acid and diethylaminoethyl methacrylate(DEAEMA).

BEST MODE FOR CARRYING OUT THE INVENTION

The invention will now be described in more detail with reference to thefollowing examples.

EXAMPLE 1 Materials

The material utilised in the examples were:

(i) hydroxyethyl methacrylate (HEMA), Methacrylic Acid (MAA), Diethylaminoethyl methacrylate (DEAEMA) and Tetraethylene glycol dimethacrylate(TEGDMA).

(ii) [¹⁴ C] Methylated Human fibronectin (specific activity 1.4Ci/mMol)and Bovine Serum Albumin (specific activity 3.5Ci/mMol)

(iii) NCS solubiliser for Liquid scintillation counting.

(iv) Bovine plasma fibronectin (FN) prepared as described in G. N.Hannan, J. W. Redmond, and B. R. McAuslan, "Similarity of thecarbohydrate moieties of fibronectins derived from blood plasma andsynthesised by cultured endothelial cells," Biochim. Biophys. Acts, 801,396 402 (1984).

(v) Virgin, unfilled P.T.F.E (TEFLON, Registered Trade Mark) sheet, 0.25mm thick

(vi) Segmented polyurethane sheet, 0.2 mm thick was cast fromcommercially available "Biomer" solution.

Preparation of polymers

PolyHEMA homopolymer was prepared essentially as described byCiverchia-Perez et. al., (L. Civerchia-Perez, B. Faris, G. LaPointe, J.Beldekas, H Leibowitz, and C. Franzblau, "Use ofcollagen-hydroxyethylmethacrylate hydrogels for cell growth," Proc.Natl. Acad. Sci. USA, 77(4), 2064-2068 (1980)) except that the castingof the hydrogels was carried out in a Bio-Rad Slab-gel pouring apparatususing 0.75 mm thick Teflon spacers. By varying the thickness of theTeflon spacers gels were successfully cast from 0.25 mm to 1 mm inthickness.

The copolymer hydrogels of polyHEMA/MAA and polyHEMA/DEAEMA wereprepared as described by Holly and Refojo (F. J. Holly, and M. F.Refojo, "Wettability of hydrogels I. Poly (2-hydroxyethylmethacrylate)," J. Biomed. mater. Res., 9, 315-326 (1975)), againcasting was done in the Bio-Rad apparatus.

After polymerisation, the hydrogels were cut into 14 mm diameter discsand dialysed extensively against phosphate buffered saline pH 7.4.(PBS)It was found that dialysing the membranes prior to cutting into discswas not as efficient with the larger sizes that were able to be cast inthe Bio-Rad system. Some swelling of the discs did occur as a result ofdialysis but they fitted readily into a 16 mm diameter tissue culturewell.

After dialysis the buttons were transferred to PBS containingpenicillin, streptomycin and kanamycin and stored at 4° C. Sterilisationby UV for 2 hours was carried out immediately prior to use for cellstudies and the sterilised discs equilibriated with an appropriatesterile cell culture medium for one hour. The teflon sheet was cut into15 mm diameter discs and the polyurethane sheet was cut into squares of1 cm×1 cm. Both materials were washed extensively with acetone and thenabsolute ethanol. Sterilisation was achieved by prolonged soaking inabsolute ethanol with washes in sterile PBS, or by autoclaving. Nodetectable differences were obvious from using either method before cellor protein binding assays.

Surface modification of polyHEMA

PolyHEMA homopolymer discs were treated individually as follows.

A disc was placed into a specially constructed polyallomer ladle (2.5 cmin diameter, 2 cm deep with a 6.5 cm×0.75 cm handle, 3 cm diameterperforations were punched out of the bottom of the ladle with a corkborer) and quickly immersed into acid (either sulphuric 98%,hydrochloric, consisting of adding 3 vol. 70% perchloric acid and 2 vol.of saturated aqueous potassium chlorate or hydrofluoric 50%) forpredetermined times ranging from 1 to 20 seconds. The perforations inthe base of the ladle allowed the acid to act equally on all hydrogelsurfaces by creating a turbulent upwelling of acid that suspended thedisc in the centre of the ladle. Gentle vertical agitation of the ladlemaintained this action. After acid immersion, the ladle and disc wereimmediately washed in deionised water several times and the disc thentransferred to PBS for extensive dialysing. The discs turned opaque infashion similar to those that were first dialysed after the initialcasting of the hydrogel, but quickly cleared. After dialysis the discswere stored and subsequently used in the same manner as described in thehydrogel preparation.

Basic hydrolysis of polyHEMA was carried out at both room temperatureand at 78° C. by immersing discs into solutions of 3.0M NaOH for timesranging from 30 minutes to 6 hours. Subsequent washing and dialysis oftreated discs was the same as described for the acid treated samples.

Cell culture and cell growth rate determination

A clonal line of normal bovine aortal endothelial cells (BAE) was grownand maintained as previously described.

Single hydrogel discs, Teflon discs and polyurethane squares were placedinto separate wells of a Costar cluster dish (24 wells, 16 mm diameter).1 ml suspensions of 5×10⁴ cells were added to each well and routinelymaintained in medium 199 plus 10% fetal calf serum.

To determine cell numbers discs were transferred from wells to 35 mmdiameter polystyrene tissue-culture dishes and fixed with 2.5%glutaraldehyde. Counts were obtained for a superimposed grid arearepresenting 0.106 mm² by using a Bioquant Image Analysis system coupledwith an Olympus BH-2 phase contrast microscope. Cell numbers are givenas the average and standard deviation of 15 random counts on each of 4discs, and are expressed per cm²

For translucent materials such as Teflon and polyurethane the cells werestained with Giemsa prior to counting.

Cell attachment determination

Cells were plated onto the various samples as described above andincubated for 6 hrs at 37° C. After incubation the cells were washedwith sterile PBS to remove those not attached. Cells on the translucentsamples were fixed and stained as described, then all samples counted onthe Bioquant System. Cell numbers are given as an average and standarddeviation of 15 random counts on each of 4 replicates per sample andexpressed as a percentage of equivalent cells attached per cm² toglow-discharge polystyrene.

EXAMPLE 2 Cell Response to Acid Etched polyHEMA

Bovine Aortal Endothelian (B.A.E.) cells were plated onto a series ofpolyHEMA substrates that had been exposed to the following sulphuricacid etching treatment times.

(a) nil

(b) 1 second

(c) 2 seconds

(d) 5 seconds

(e) 10 seconds

(f) 15 seconds

The percent cell attachment as a function of the sulphuric acid etchingtime for the polyHEMA substrates is shown in FIG. 1. The percent cellattachment for hydrochloric acid and hydrofluoric acid etched polyHEMAsubstrates is also shown in FIG. 1 to emphasise the efficacy of thesulphuric acid etch.

EXAMPLE 3 Relative Placing Efficiencies of B.A.E. Cells on EtchedpolyHEMA v. Teflon

Bovine Aortal Endothelial (B.A.E.) cells were plated onto a sulphuricacid etched polyHEMA substrate and onto a P.T.F.E. (TEFLON) substrateand the number of cells attached after six hours was expressed as apercentage of the number that attached to tissue culture polystyrene Theresults were:

TEFLON 70-72%

Etched PolyHEMA: 90-95%

The sulphuric acid etched polyHEMA was relatively more efficient in itsplating efficiency with respect to tissue culture polystyrene than theP.T.F.E.

EXAMPLE 4 Comparative Cell Growth of B.A.E. on Etched polyHEMA andTissue Culture Polystyrene

The growth of B.A.E. cells (number per square millimeter) at 3, 6 and 10days is shown in FIG. 2 for sulphuric acid etched polyHEMA andglow-discharge treated polystyrene. B.A.E. cell growth to confluence onetched polyHEMA was comparable to that on tissue culture polystyreneboth in terms of growth rate and morphology.

EXAMPLE 5 Platelet Binding

Human platelets were prepared from fresh blood and labelled with ⁵¹ Cr(spec. act. 635 uCi/ml) essentially as described by Dacie and Lewis (J.V. Dacie, and S. M. Lewis, Practical Haematology, 5th. edn., ChurchillLivingstone, Edinburgh (1975). Samples of polymers were mixed withlabelled platelets (conc. 5.0×10⁸ /ml) and agitated gently for 2 hrs.After mixing the polymers were washed three times with PBS containing 1%BSA then counted. The cell numbers of platelets remaining bound wereexpressed per cm² of sample.

Because the application of an endothelial cell binding material tovascular prostheses is ultimately linked to the problem ofthrombogenicity, at least a preliminary indication of the thrombogenicpotential of a material might be gained from its propensity for plateletbinding. From the results obtained (Table III) it is clear that theaffinity of platelets for "etched" pHEMA was much greater than forsegmented polyurethane or any of the other polymers tested.

EXAMPLE 6 Protein Binding

Materials to be tested were placed individually into Costar cluster dishwells. 0.5 ml of sterile PBS was added to each well, then an appropriatealiquot of either [¹⁴ C] methylated Human fibronectin or [¹⁴ C]methylated bovine serum albumin added to give a final concentration of0.1 uCi per well. The wells were incubated for 45 mins at 37° C. Afterincubation the materials were transferred to 20 ml of fresh PBS andwashed for one hour, a further rinse in 5 ml of PBS was carried outprior to transferring the materials to liquid scintillation vials. 4 mlof NCS tissue-solubilising solution was added to each vial and then thevials were incubated at 5° C. for 2 hours.

After cooling to room temperature, an appropriate scintillant was addedand the samples dark equilibriated at 4° C. overnight prior to counting.The amount of fibronectin and bovine serum albumin bound is the averageobtained from 3 buttons per sample and expressed as a percentage of thatbound per cm², to glow-discharge treated polystyrene tissue culturedishes.

Initial studies on the propensity of sulphuric acid treated ("etched")polyHEMA to bind blood proteins were conducted with bovine serum albuminand fibronectin. For comparison we used T/C polysty, Teflon and asegmented polyurethane (Biomer). The results are presented in Table IIand FIG. 3. PolyHEMA was remarkable in that it showed very weak bindingof albumin compared with the other polymers. Acid "etching" did notchange this quantitatively. It is known that pHEMA does not bindfibronectin, a major adhesive component of the extracellular matrix.Although acid "etching" did bring about a measurable change, theresultant fibronectin binding capacity was of the order of 10 fold lessthan for the other polymers studied.

Electron spectroscopy for chemical analysis (ESCA) was used to determinechemical changes in the polymer surfaces. Replicate samples of polyHEMAhydrogel treated with either chloric, hydrofluoric or sulphuric acidswere prepared. Samples of each were assayed to ensure their celladhesive properties were as expected and parallel samples were submittedfor ESCA analysis.

For each acid used the most obvious alteration to the polyHEMA surfacewas indicated by spectral shifts. These changes indicated a significantchemical modification consistent with the creation of surface --COOHgroups. Problems raised regarding the correlation of apparent surfacechanges to biological responses are discussed below.

EXAMPLE 7 Endothelial Cell Attachment to Copolymer Hydrogels

Very little is known about the specific molecular requirements forendothelial cell adhesion in vascular prostheses. The polyHEMA hydrogelwith its neutrally charged hydroxyl rich surface is a potentially usefulsystem to explore the requirements for cell attachment, but it has beenestablished that mammalian cells will not adhere to and grow onhydrogels of polyHEMA homopolymer.

To follow the effects of introducing charged groups into the polyHEMAsurface a series of hydrogels were prepared from copolymers of HEMA withincreasing amounts of methacrylic acid (MAA) or HEMA with increasingamounts of diethylaminoethyl methacrylate (DEAEMA), thus introducingnegative carboxyl (--COOH) or positive (amino) charges respectively.

Using endothelial cell attachment (and subsequent growth) as parameters,these copolymers were compared to glow-discharge treated tissue culturegrade polystyrene. The attachment capacity of polystyrene forendothelial cells was set at 100% for comparison. When compared withpolystyrene the attachment of vascular endothelial cells determined as aplating efficiency, was increased from negligible levels on polyHEMAhomopolymer to levels of about 50% after copolymerisation with eitherMAA or DEAEMA (FIG. 4). Optimal levels were 20% DEAEMA or 30% MAA v/v.Beyond these levels DEAEMA inclusion caused cytotoxic effects leading tocell detachment and arrest of growth.

Methacrylic acid inclusion caused no deleterious effect on cellmorphology or growth, but beyond addition to 30% v/v the hydrogel becamefriable and so physically altered as to be impractical to handle.

The above results discussed in relation to Examples 2 to 7 suggestedthat either positive or negative charge groups could affect endothelialcell adhesion. The introduction of carboxyl groups was focused on aswere other means of introducing such surface charges. The change inproperties of polystyrene by acid oxidation is well documented and wetried a similar approach with polyHEMA. PolyHEMA hydrogel buttons weretreated with either sulphuric acid hydrofluoric acid or chloric acid.The latter has been shown to be highly effective in creating a celladhesive surface on polystyrene. The buttons treated for varying timeswere subsequently washed free of acid then tested for ability to supportendothelial cell attachment and growth.

Treatment with either chloric or hydrofluoric acids over a wide range oftimes caused no demonstrable change in the cell adhesion properties ofpolyHEMA. In contrast sulphuric acid treatment profoundly altered thesurface of the hydrogel such that it became excellent for the adhesionand spreading of vascular endothelial cells (FIG. 1). The morphologicappearance of cells, 24 hours after seeding onto sulphuric acid etchedpolyHEMA, is shown (FIG. 2) and, provided the optimal etch time (10 sec)was not exceeded, was indistinguishable from those grown on tissueculture grade polystyrene (T/C polysty) The efficiency of attachment ofcells to sulphuric acid treated polyHEMA was compared also to Teflon andsegmented polyurethane (Table I). The results suggest that etchedpolyHEMA in this respect was practically as effective as T/C polysty andbetter than Teflon or polyurethane.

EXAMPLE 8 Endothelial Cell Attachment to Alkali-Treated polyHEMA

In order to determine if cell attachment to surface modified polyHEMAcould be induced by non-acid means, several discs were subjected to basehydrolysis. No obvious physical change to the surface of the discs wasnoted in any of the treatments with 3.0M NaOH at room temperature. Thosediscs subjected to increasing times in 3.0M NaOH at 78° C., however,showed signs of surface cracking after 6 hours (that was similar to a 15second treatment with sulphuric acid). Subsequent testing of all thealkali treated discs failed to reveal any changes in the ability of thehydrogel to support cell attachment and growth.

EXAMPLE 9 Cell Growth Rate on Acid-Treaded polyHEMA

Sulphuric acid-treated polyHEMA hydrogel was compared to T/C polystyrenefor their ability to support growth of aortal endothelial cells (BAE).No significant difference between them was found in respect to rate ofcell growth and final cell density achieved.

The failure of polyHEMA to support adherence of mammalian cells is welldocumented. A contrary report often cited in reference to cell growthcontrol has been dismissed as an artefact of discontinuouscell-substratum interactions. The results show that brief exposure of apolyHEMA hydrogel surface to concentrated sulphuric acid results in asubstratum to which vascular endothelial cells attach and grow virtuallyas well as they do on the best available tissue culture grade ofglow-discharge treated polystyrene. Preliminary results indicate that apolyHEMA. The "etched" polyHEMA surface gave a more uniform attachmentand subsequent growth of endothelial cells than did Teflon and this maybe advantageous where preliminary endothelial cell seeding can beemployed and where uniform surface repopulation is desirable.

Early work on polystyrene "etching" first centred on sulphonic groupsfor cell attachment. This idea was subsequently discounted and laterstudies concluded that introduction of --OH groups rather than --COOHgroups were essential for mammalian cell attachment. This does notappear likely in the polyHEMA system where it is initially an hydroxylrich surface that is rendered adhesive following the introduction of--COOH groups.

ESCA analysis of the sulphuric acid treated polyHEMA indicated that themajor change was due to creation of --COOH groups which would haveresulted in an increase in negative charges on the hydrogel surface.This was confirmed by observations on a corresponding increase in theaffinity for cationic dyes (e.g. Crystal Violet, Acridine Orange). Thisanalysis also correlates with the enhanced cell attachment propertyintroduced by copolymerisation with methacrylic acid.

Preliminary evidence suggests that glycol was a by-product of the"etching" procedure. Thus we propose that a likely effect of briefsulphuric acid "etching" is to enhance cell interaction by a limitedhydrolysis of polyHEMA to produce polymethacrylic acid as shown:##STR1## Methacrylic acid groups on the hydrogel surface may alsosufficiently alter the degree of hydration to promote cell attachment.

Hydrochloric or hydrofluoric acid "etching" lead to reformation ofsurface carboxyl groups and surfaces which presented similar spectra tothat elicited by sulphuric acid "etching". However, hydrochloric orhydrofluoric treatment consistently failed to change the adhesivecharacteristics of polyHEMA for cells. The hydrolytic treatment ofpolyHEMA under alkaline conditions, however, demonstrates that simplehydrolysis by acid or basic means elicits changes that do notnecessarily lead to conditions suitable for cell attachment. Conceivablyhigher cross-linking or specific--COOH orientation may occur with thosetreatments that fail to produce such surface modification.

It is conceivable that attachment of cells to "etched" polyHEMA ismediated by a serum protein such as fibronectin The small butsignificant increase in ability of polyHEMA to bind fibronectinfollowing acid "etching" is not inconsistent with this idea. Howeverhydrochloric acid or hydrofluoric acid "etching" hydrogels boundfibronectin just as efficiently (Table II) but these did not supportcell attachment. Therefore it is unlikely that cell attachment tosulphuric "etched" polyHEMA is simply a result of its ability to bindfibronectin.

Various modifications may be made in details of composition and of theprocess without departing from the scope and ambit of the invention.

                  TABLE I                                                         ______________________________________                                                       Cell Attachment (6 hr)                                         Polymer        % per cm.sup.2 *                                               ______________________________________                                        T/C polysty    100                                                            Etched polyHEMA                                                                              90                                                             Teflon         75                                                             Polyurethane   73                                                             polyHEMA       1                                                              ______________________________________                                         *T/C polysty set as maximal attachment.                                  

Relative attachment of BAE cells to various polymers. Numbers attachedwere expressed as a percentage of the number of cells attaching to T/Cpolysty.

                  TABLE II                                                        ______________________________________                                        FN and BSA Binding                                                                       FN          BSA                                                               pmol · cm.sup.2                                                              %       pmol · cm.sup.2                                                                %                                        ______________________________________                                        polysty      1.31      100     1.20    100                                    polyHEMA     0         0       0.03    2.5                                    etched polyHEMA                                                                            0.21      16.0    0.03    2.5                                    (H.sub.2 SO.sub.4)                                                            etched polyHEMA                                                                            0.14      10.7    0.02    1.7                                    (Chloric)                                                                     Teflon (P.T.F.E.)                                                                          1.74      132.8   1.88    156.7                                  polyurethane (Biomer)                                                                      1.34      102.3   2.03    169.2                                  ______________________________________                                         Binding of fibronectin (FN) and bovine serum albumin (BSA) to polymers.  

                  TABLE III                                                       ______________________________________                                        Relative Platelet Binding                                                                      No. Platelets                                                Polymer          Bound/cm.sup.2                                               ______________________________________                                        polyHEMA         1.0 × 10.sup.5                                         polyHEMA etched  2.0 × 10.sup.6                                         P.T.F.E. (Teflon)                                                                              6.8 × 10.sup.5                                         polyurethane (biomor)                                                                          6.5 × 10.sup.5                                         ______________________________________                                    

Human platelets prepared from fresh blood were labelled with ⁵¹ Cr.Labelled platelets were incubated with polymer samples and number ofplatelets bound per cm² in 12 hours from the known specific activity ofthe inoculum and the radioactivity bound to samples.

I claim:
 1. An implantable material comprising a substrate having asurface and a polyHEMA hydrogel applied to said surface which ischemically modified by means of hydrolytic etching by an acid in orderto alter the condition of said surface form a non-adhesive state to anadhesive state and thereby promote the attachment and growth of cells tosaid surface.
 2. An implantable material according to claim 1 whereinthe hydrogel is a synthetic hydrogel.
 3. An implantable materialaccording to claim 1 wherein said acid is sulphuric acid of strength 36N(98%).
 4. An implantable material according to claim 1 wherein saidhydrolytic etching is carried out for a period of less than 15 seconds.5. A method of forming an implantable material comprising the stepsof:(a) forming a substrate having a surface, (b) applying a polyHEMAhydrogel layer to the surface of the substrate, and (c) chemicallymodifying the polyHEMA hydrogel by means of hydrolytic etching by anacid in order to alter the condition of said surface from a non-adhesivestate to an adhesive state and thereby promote the attachment and growthof cells to said surface.
 6. A method according to claim 5 wherehydrolytic etching by an acid utilizes sulphuric acid of a strength 36N(98%).
 7. A method according to claim 6 wherein said acid etching isperformed for a period of less than 15 seconds.